Abstract Q fever is a worldwide zoonotic disease that is caused by the obligate intracellular Gram-negative bacterium, Coxiella burnetii. Human Q fever can develop into a severe chronic, potentially fatal disease. Although this organism previously weaponized and currently classified as a category B select agent, no vaccine is commercially available for the prevention of human Q fever in the US. Therefore, the development of safe and effective prophylactic and therapeutic strategies for controlling C. burnetii infection remains an important goal for public health and national biosecurity. However, there is a fundamental gap in the knowledge regarding the mechanisms of host immune response to C. burnetii natural infection. To fill this gap, this new R21 application aims to explore whether B1 B cells play a critical role in controlling bacterial replication and regulating host immune response during C. burentii aerosol infection. Despite C. burnetii being an intracellular bacterial pathogen, our preliminary study demonstrates that a C. burnetii infection induces more severe splenomegaly and higher bacterial burden in the spleens of B1 cell deficient mice, suggesting that B1 cells play an important role in host defense against primary C. burnetii infection. The objective of this application is to use a C. burnetii aerosol infection mouse model to determine the mechanisms of B1 cell-mediated protective immunity against C. burnetii natural infection. To achieve this objective, two specific aims were proposed to test the central hypothesis that B1 cell-mediated protective immunity against C. burnetii infection depends on its abilities to control bacterial replication and regulate inflammatory response. Aim 1 will determine if B1 cells can mediate killing of C. burnetii. We will determine whether i) B1 cells play a critical role in host defense against C. burnetii aerosol infection; and ii) B1 cells can function as phagocytes to mediate clearance of C. burnetii. Aim 2 will determine if B1 cells can regulate C. burnetii infection-induced inflammatory responses. We will determine i) whether B1 cells are crucial for regulating aerosolized C. burnetii-induced inflammatory response; and ii) how B1 cells regulate inflammatory response during C. burnetii infection. As an outcome of the proposed research, we expect to enhance our understanding of the mechanisms of B1 cell-mediated protective immunity against C. burnetii natural infection. This will have significant positive effects on human health, because it will provide critical information for developing novel immunotherapeutic strategies against Q fever.